Neurodegenerative disorders such as Parkinson's, Alzheimer's, and Huntington's disease are becoming ever more prominent in our society. A direct approach towards therapeutic treatment of these diseases is through replacement therapy where undamaged tissue is transplanted into the nervous system. Recently, significant progress has been achieved with transplants in Parkinson's disease (PD), but the process is heavily dependent on an unstable and problematic source of fetal tissue. Embryonic stem (ES) cells may be a tissue/cell source for developing the therapeutic potential of neural transplantation. Self-renewing and multipotent stem cells provide a source of transplantable material to replace post-mitotic neurons that do not spontaneously regenerate after injury.
PD is a progressive neurodegenerative disease characterized clinically by bradykinesia, rigidity, and resting tremor. The motor abnormalities are associated with a specific loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) and depletion of striatal dopamine (DA) levels. While the loss of striatal DA correlates with the severity of clinical disability, clinical manifestations of PD are not apparent until about 80-85% of SNc neurons have degenerated and striatal DA levels are depleted by about 60-80%. DA neurons in the ventral midbrain consist of two main groups: the A9 group in the SN, and the A10 group in the medial and ventral tegmentum. Each of these cell groups project to different anatomical structures and is involved in distinct functions. A9 cells mainly project to the dorsolateral striatum, involved in the control of motor functions, whereas A10 cells provide connections to the ventromedial striatum, limbic and cortical regions, involved in reward and emotional behavior. In addition to the distinct axonal projections and differences in synaptic connectivity, these groups of DA cells exhibit differences in neurochemistry and electrophysiological properties, illustrating functional differences despite similar neurotransmitter identity. These differences in A9 and A10 cells are also reflected in their specific responses to neurodegeneration in PD. Postmortem analyses in human PD brains demonstrate a selective cell loss of the A9 group with a survival rate of about 10% whereas the A10 group is largely spared with a survival rate of about 60%. This indicates that A9 cells are more vulnerable to intrinsic and/or extrinsic factors causing degeneration in PD. In addition, three regional gradients of neurodegeneration in the dorso-ventral/rostro-caudal/medio-lateral axis have been reported in PD. Caudally and laterally located ventral DA cells within A9 subgroups are the most vulnerable cells in PD. In contrast, the medial and rostral part of DA cell subgroups within A10 cells (i.e. rostral linear nucleus, RLi) are the least affected (5-25% cell loss).
Current clinical data indicate proof of principle for cell implantation as a therapy for PD. It has recently been shown that ES cells, when transplanted in low numbers into the striatum, develop into fully-differentiated DA neurons that can restore cerebral function and behavior in an animal model of PD (Björklund et al., Proc. Natl. Acad. Sci. USA 99:2344-2349, 2002). Furthermore, the PD process does not appear to negatively affect the transplanted cells; however, the endogenous DA neurons continue to degenerate. The stem cells may themselves be transplanted or, alternatively, they may be induced to produce differentiated cells (e.g., neurons, oligodendrocytes, Schwann cells, or astrocytes) for transplantation.
Differentiation of stem cells into various cell types is regulated by a host of intracellular and extracellular factors. For example, the transforming growth factor-β (TGF-β) superfamily of secreted polypeptide factors regulates a variety of homeostatic and developmental processes, including stem cell determination and differentiation during embryogenesis. A basic understanding of the signal transduction pathway from the TGF-β signal to the target genes that generate the biological responses has been worked out (Attisano et al., Science 296:1646-1647, 2002). Binding of a TGF-β member to an appropriate cell-surface receptor leads to phosphorylation of intracellular protein mediators called Smads, in particular the class of Smads known as the R-Smads, which then accumulate in the nucleus as heteromeric complexes with a second class of Smads, the Co-Smads (of which Smad4 is the only known mammalian example at this time). In the nucleus, the R-Smad/Co-Smad complexes associate with particular transcriptional coactivators or corepressors, thereby differentially affecting gene expression and generating diverse biological responses. Inhibitory Smads, or I-Smads (e.g., Smad6 and -7), which antagonize TGF-β signaling, are also known.
Included in the TGF-β superfamily are nodal, bone morphogenic proteins (BMPs), and activins. Mice homozygous for a mutation of the nodal gene fail to form the primitive streak and most of the mesoderm. Pathway for transducing the nodal signal involves activin receptor-like kinase receptors (e.g., ALK4 and ALK7) and SMADs (e.g., Smad2), and is regulated by members of the ECF-CFC family (e.g., Cryptic, Oep (from zebrafish), and Cripto). Similarly, activins, which induce mesodermal tissues and endoderm, operate through signal transduction pathways that involve receptors (e.g., activin receptor I, activin receptor II, activin receptor IIb, TGF-β receptor, ALK4, and ALK7) and SMADs (e.g., smad2, smad3, and smad4). Related to activin, BMP (e.g., BMP2, BMP4, and BMP7) utilizes signal transduction pathways that involve BMP receptors (BMPR) and activin receptors (e.g., BMPRIa, BMPRIb, BMPRII, ALK1, ALK2, ALK3, and ALK6), and SMAD proteins (e.g., Smad4, Smad5, and Smad6).